LOCUS       pTW086M_Lvl0_mKa        3424 bp    DNA     circular     08-APR-2014
DEFINITION  No BsaI or SapI.
ACCESSION   urn.local...k2-9e4hqdu
KEYWORDS    .
SOURCE      Cloning vector pUC19
  ORGANISM  Cloning vector pUC19
            other sequences; artificial sequences; vectors.
REFERENCE   1  (bases 1 to 2686)
  AUTHORS   New, England and Biolabs.
  TITLE     Direct Submission
  JOURNAL   Submitted (18-OCT-2007) Research Department, New England Biolabs,
            240 County Road, Ipswich, MA 01938, USA COMMENT See also GenBank
            accession L09137.
FEATURES             Location/Qualifiers
     source          1..3424
                     /organism="Cloning vector pUC19"
                     /mol_type="other DNA"
     gene            55..655
                     /standard_name="lacZ PCR Product from pUC19"
     primer_bind     55..75
                     /standard_name="TK_DI_lacZ_Fwd_1"
     CDS             complement(146..469)
                     /gene="lacZalpha"
                     /codon_start=1
                     /product="beta-galactosidase alpha fragment"
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLN
                     RLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTI
                     CSDAA"
     gene            complement(146..469)
                     /gene="lacZalpha"
     primer_bind     364..386
                     /standard_name="M13/pUC Forward (Quintara)"
     primer_bind     378..395
                     /standard_name="M13 (-21) Forward (Quintara)"
     misc_feature    396..452
                     /gene="lacZalpha"
                     /note="multiple cloning site (EcoRI-HindIII)"
                     /note="Geneious type: polylinker"
     primer_bind     complement(465..481)
                     /standard_name="M13 Reverse (Quintara)"
     -10_signal      complement(514..519)
                     /note="Plac promoter (drives lacZalpha;
                     counter-clockwise); TATGTT"
     -35_signal      complement(538..543)
                     /note="Plac promoter (drives lacZalpha;
                     counter-clockwise); TTTACA"
     protein_bind    complement(563..575)
                     /note="CAP protein binding site"
                     /bound_moiety="catabolite activator protein (CAP)"
     primer_bind     complement(637..655)
                     /standard_name="TK_DI_lacZ_Rev_1"
     misc_feature    679..693
                     /standard_name="15 bp SapI Overhang"
     primer_bind     679..726
                     /standard_name="TW_lvl0_mKate_SapI_F"
     primer_bind     690..726
                     /standard_name="TW_lvl0_mKate_IF_Fwd"
     primer_bind     690..704
                     /standard_name="15 bp for In-Fusion"
     misc_feature    694..699
                     /standard_name="BsaI"
     primer_bind     694..726
                     /standard_name="TW_lvl0_mKate_Fwd1"
     primer_bind     694..721
                     /standard_name="TW_lvl0_mKate_Fwd"
     misc_feature    701..704
                     /standard_name="BsaI Cut Site"
     primer_bind     701..721
                     /standard_name="mKate Binding"
     CDS             705..1418
                     /standard_name="mKate"
     primer_bind     705..726
                     /standard_name="mKate binding 2"
     misc_feature    722
                     /note="Geneious type: Editing History Replacement"
                     /standard_name="A"
     primer_bind     complement(1005..1024)
                     /standard_name="mKate-seq-R"
     misc_feature    1343
                     /note="Geneious type: Editing History Replacement"
                     /standard_name="C"
     primer_bind     1380..1400
                     /standard_name="TW_mKate-HDV-seq-F"
     primer_bind     complement(1393..1443)
                     /standard_name="TW_lvl0_mKate_SapI_F"
     primer_bind     complement(1393..1432)
                     /standard_name="TW_lvl0_mKate_IF_Rev"
     primer_bind     complement(1393..1428)
                     /standard_name="TW_lvl0_mKate-Rev1"
     primer_bind     complement(1397..1428)
                     /standard_name="TW_lvl0_mKate_Rev"
     primer_bind     complement(1397..1418)
                     /standard_name="mKate binding"
     gene            1418
                     /standard_name="mVenus"
     misc_feature    complement(1418..1421)
                     /standard_name="BsaI Cut Site"
     primer_bind     complement(1418..1432)
                     /standard_name="15 bp In-Fusion"
     misc_feature    complement(1423..1428)
                     /standard_name="BsaI"
     primer_bind     1429..1443
                     /standard_name="15 bp SapI Overhang"
     rep_origin      complement(1546..2219)
     misc_RNA        complement(1605..2157)
                     /note="RNAII transcript (complementary strand)"
     rep_origin      complement(1605..2193)
     rep_origin      complement(1605..2138)
     primer_bind     complement(1644..1664)
                     /standard_name="TW_HDV-seq-R"
     primer_bind     1645..1666
                     /standard_name="pHAGE-A7550-7050"
     primer_bind     complement(1909..1933)
                     /standard_name="pHAGE-S7501-8000"
     primer_bind     complement(1953..1972)
                     /standard_name="RepO-R"
     -35_signal      2011..2016
                     /note="RNAI promoter (clockwise); TTGAAG"
     -10_signal      2033..2038
                     /note="RNAI promoter (clockwise); GCTACA"
     misc_RNA        2047..2154
                     /note="RNAI transcript"
     primer_bind     2156..2177
                     /standard_name="pHAGE-A7050-6550"
     -10_signal      complement(2167..2172)
                     /note="RNAII promoter (counter-clockwise); CGTAAT"
     -35_signal      complement(2188..2193)
                     /note="RNAII promoter (counter-clockwise); TTGAGA"
     CDS             complement(2364..3224)
                     /gene="bla"
                     /note="ampR (confers resistance to ampicillin)"
                     /codon_start=1
                     /product="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY
                     IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVE
                     YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL
                     DRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL
                     LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA
                     EIGASLIKHW"
     gene            complement(2364..3224)
                     /gene="bla"
     primer_bind     complement(2413..2437)
                     /standard_name="pHAGE-S7001-7500"
     misc_feature    complement(2504..2509)
                     /standard_name="BsaI site"
     primer_bind     2665..2688
                     /standard_name="pHAGE-A6550-6050"
     primer_bind     complement(2933..2957)
                     /standard_name="pHAGE-S6501-7000"
     primer_bind     2987..3006
                     /standard_name="Amp-R; Quintara AmpR"
     primer_bind     3134..3157
                     /standard_name="pHAGE-A6050-5550"
     sig_peptide     complement(3156..3224)
                     /gene="bla"
                     /note="required for secretion to the periplasm; cleaved
                     off to form the mature beta-lactamase protein."
     promoter        complement(3225..3259)
                     /note="Beta Lactamase Promoter"
                     /standard_name="P-Bla"
     -10_signal      complement(3268..3273)
                     /note="bla promoter (counter-clockwise); GAGACA"
     -35_signal      complement(3289..3294)
                     /note="bla promoter (counter-clockwise); TTCAAA"
ORIGIN      
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt cgagctcggt acccggggat
      421 cctctagagt cgacctgcag gcatgcaagc ttggcgtaat catggtcata gctgtttcct
      481 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
      541 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
      601 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
      661 agaggcggtt tgcgtattgg gcgctcttcc gctggtctca caccatggtg tctaagggcg
      721 aggagctgat taaggagaac atgcacatga agctgtacat ggagggcacc gtgaacaacc
      781 accacttcaa gtgcacatcc gagggcgaag gcaagcccta cgagggcacc cagaccatga
      841 gaatcaaggt ggtcgagggc ggccctctcc ccttcgcctt cgacatcctg gctaccagct
      901 tcatgtacgg cagcaaaacc ttcatcaacc acacccaggg catccccgac ttctttaagc
      961 agtccttccc tgagggcttc acatgggaga gagtcaccac atacgaagac gggggcgtgc
     1021 tgaccgctac ccaggacacc agcctccagg acggctgcct catctacaac gtcaagatca
     1081 gaggggtgaa cttcccatcc aacggccctg tgatgcagaa gaaaacactc ggctgggagg
     1141 cctccaccga gatgctgtac cccgctgacg gcggcctgga aggcagaagc gacatggccc
     1201 tgaagctcgt gggcgggggc cacctgatct gcaacttgaa gaccacatac agatccaaga
     1261 aacccgctaa gaacctcaag atgcccggcg tctactatgt ggacagaaga ctggaaagaa
     1321 tcaaggaggc cgacaaagag acatacgtcg agcagcacga ggtggctgtg gccagatact
     1381 gcgacctccc tagcaaactg gggcacaaac ttaattgata atgagaccgc ttcctcgctc
     1441 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     1501 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     1561 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     1621 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     1681 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     1741 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     1801 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     1861 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     1921 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     1981 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     2041 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     2101 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     2161 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     2221 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     2281 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     2341 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     2401 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     2461 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     2521 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     2581 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     2641 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     2701 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     2761 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     2821 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     2881 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     2941 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     3001 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     3061 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     3121 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     3181 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     3241 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     3301 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
     3361 taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt
     3421 cgtc
//
